Introduction to Plant Virology History Definitions Classification Structure Additional program (students presentation): Mimivirus (Raoult et al.
al., 2004, SCIENCE 306, p. 1344 ff, Xiao et al., J. Mol. Biol. (2005) 353, 493496)
K. Richert-Pggeler, WS 05/06
Roger Hull: Matthews Plant Virology (2nd edition, 2004)
[Link] [Link]
First plant virus description 752 AD
Eupatorium yellow-vein (gemini) virus (EpYVV)
Saunders et al., 2003
Dr Robert G. Milne, CNR, Instituto di Fitovirologica Applicata, Torino, Italy
Tulip breaking (Poty) virus
Ambrosio Bosschaert, Dutch painter, 1573-1621
Brunt and Walsh, 2005
1892: Dmitri Iwanowski, Russian scientist, works with tobacco plants with Tobacco Mosaic Disease -discovers filtration does not remove infectious agent for TM disease -could not visualize agent with microscope, nor grow on microbial media -concluded he had found infectious agent smaller than bacterium
infection is not caused by microbes but by a contagium vivum fluidum reproduces itself in the diseased plants other diseases of unknown cause may be ascribed to a contagium fluidum
VIRUS (lat.): venom, slime
Martinus W. Beijerinck (1851-1931) ber ein contagium vivum fluidum als Ursache der Fleckenkrankheit der Tabaksbltter. Verhandel. Acad. Wetensch. Amsterdam 65:3-21 (1898)
John Shaw, 2002
Kochs Postulates
1843-1910, Nobel Prize 1905
1. The causal agent must be associated in every case with the disease as it occurs naturally. 2. The causal agent must be isolated in pure culture 3. When the host is inoculated the characteristic symptoms of the disease must develop. 4. The causal agent must be reisolated.
Visualization of viruses made possible by electron microscope
Magnification up to 106 fold !!!! nm=10-9 meter
The original electron microscope as developed in 1938 in McLennan Laboratories of the University of Toronto is now on permanent exhibition at the Ontario Science Centre, Toronto, Ontario
Isolation of a crystalline protein possessing the properties of tobacco mosaic virus. Science 81:644-645, 1935
Wendell Stanley, 1904-1971
By courtesy of the Molecular Biology & Virus Laboratory, University of California, Berkeley
Stanley achieved the first crystallization of a virus (1935), the basis for his Nobel Prize of 1946 (Chemistry). He later remarked on the unique position of viruses at the junction of life and non-life: "The fact that, with respect to size, the viruses overlapped with the organisms of the biologist at one extreme and with the molecules of the chemist at the other extreme only served to heighten the mystery regarding the nature of viruses. Then too, it became obvious that a sharp line dividing living from non-living things could not be drawn and this fact served to add fuel for discussion of the age-old question of 'What is life?'"
Molecular Biology and Biotechnology 1969-Restriction endonuclease cloned (Arber & Smith) 1970-Reverse transcriptase (Temin & Baltimore) 1973-Recombinant plasmid (Cohen & Boyer) 1977-DNA sequencing (Gilbert & Sanger)
1977: bacteriophage (ssDNA), 1980: CaMV (dsDNA), 1982: TMV (ssRNA)
1984-Polymerase chain reaction (PCR) (Mullis) 1995-Entire genome sequenced (Haemophilus influenzae)
Distinguishing viruses from other organisms
What fails to distinguish viruses from cellular organisms? 1. Size
The Mimivirus has a size larger than the smallest bacteria and, with about 900 genes, a genetic complexity greater than that of the most reduced bacteria
[Link]
Matthews, R. E. F. (1981). Plant Virology. Academic Press. Closteroviridae: 1.9x104 Nanoviridae: 1x103 Mimiviridae (amoeba):1x106
Distinguishing viruses from other organisms
What fails to distinguish viruses from cellular organisms? 2. Obligate intracellular parasite
Fails to distinguish viruses from many bacteria, Mycoplasma, Rickettsiae and Chlamydiae
Mycoplasma: 150-300 nm diamenter, bilayer membrane, ribosomes, DNA no cell wall. Replication by binary fission. Rickettsiae: nonmotile bacteria (typhus fever), CW, plasmamembrane, ribosomes, DNA, binary fission, ATP production. Chlamydiae: psittacosis, elementary-, reticulate bodies (bilayer membrane, binary fission)
3. Stable, inert phase in life cycle
Many bacterial spores are more stable than some virus particles
General characteristics of viruses
A. Acellular, dont synthesize a cell membrane (+/- envelope= stolen host cell membrane) B. Genome = RNA or DNA C. Protein coat = capsid D. No ribosomes. Lack ability to synthesize organic molecules E. No metabolism. Cant generate own energy therefore are metabolic parasites F. Obligate intracellular parasites-can only replicate inside another host cell G. Host cell specificity: all cellular organisms may be attacked 1. Viral adhesins must bind specific host cell surface receptors 2. Appropriate host enzymes for viral replication 3. Ability of replicated viruses to be released from host cell H. Viruses do not grow, nor divide. Viruses direct synthesis of viral nucleic acid and viral proteins by host cell. Viruses are assembled.
Distinguishing virion from virus
Virion the particle that is the extracellular phase of the infection cycle, typically composed of the genomic nucleic acid and coat protein but may have a lipid membrane and other components. Intact non-replicating virus particles, no signs of life Virus virion plus intracellular aspects, including replication intermediates
Alive? viruses reproduce; property of life occur as populations have variation that is inherited
Why viruses are non-living Lack a complete protein synthesis system Lack a complete energy generation system
Virus Disease Symptoms
Local lesions Stunting
Barley yellow dwarf virus
Yellowing
Beet mild yellowing virus
Ringspot
PV-Y
Necrosis
Mosaic
Abutilon mosaic virus
Tomato spotted wilt virus
Developmental abnormalities
Tobacco mosaic virus Alfalfa mosaic virus Zucchini yellow mosaic virus
Microsymptoms
Chloroplast Degeneration (tymoviruses) Enlarged Nuclei (rhabdoviruses) Disorganized Mitochondria (aggregation (potyvirus), modification (tombusvirus) Inclusion Bodies (caulimoviruses, potyviruses)
Cytoplasmic inclusion bodies
Pinwheel Inclusions (Potyvirus) Inclusion bodies (caulimovirus)
Originate and develop in association with the plasma membrane CI protein of potyviruses RNA replication
Lesemann and Casper, 1973, Phytopathology 63
Protein enoded by gene 6 of CaMV Host range Symptom expression Translation (transactivator) DNA replication
Symptoms are not sufficient to classify virus:
mixed infections distinct strains that cause different symptoms in same host same symptoms, but different viruses:
Tobacco mosaic virus (ssRNA) Cauliflower mosaic virus (dsDNA) Abutilon mosaic virus (ssDNA)
mixed infected petunia (Richert, 1992): PVCV, CMV, PV-Y, TMV
Virus classification
Nucleic acid Morphology Genome organisation Transmission vector
[Link]
Entrez Genomes currently contains 2139 Reference Sequences for 1486 viral genomes and 36 Reference Sequences for viroids. Deltavirus dsDNA viruses, no RNA stage ssRNA negativestrand viruses unclassified viruses Retro-transcribing viruses dsRNA viruses ssRNA positive-strand viruses, no DNA stage Satellites ssDNA viruses unclassified bacteriophages
Comments and suggestions to: [genomes@[Link]] Revised: October 18, 2005
Comparitive abundance of different viruses
+ssRNA
600
dsDNA -ssRNA dsRNA
300
ssDNA rtDNA
H. Scholthof
Relative abundance of different plant viruses
+ssRNA
dsDNA -ssRNA dsRNA
ssDNA
rtDNA
H. Scholthof
Classification of plant viruses
Genome (DNA or RNA)
dsDNA-RT Caulimoviridae (pararetroviruses, vertebrates) ssDNA Geminiviridae Nanoviridae
dsRNA Partitiviridae (fungi) Reoviridae (invertebrates, vertebrates, fungi)
ssRNA(-) Rhabdoviridae (invertebrates, vertebrates) Bunyaviridae (invertebrates, vertebrates) Tenuivirus, Varicosavirus
ssRNA-RT Pseudoviridae (invertebrates, fungi) Metaviridae (invertebrates, fungi) ssRNA(+) Bromoviridae*, Comoviridae, Sesquiviridae, Tombusviridae, Luteoviridae, Tymoviridae Flexivirdae, Potyviridae#, Closteroviridae Ourmiavirus Tobamovirus, Tobravirus, Hordeivirus, Benyvirus, Pomovirus, Furovirus Pecluvirus * Alphavirus, # Picornaviridae
Virus Particle Structure
Bacilliform
Flexuous rod
Geminate
Spherical
Rigid rod
Morphology
COMPOSITION OF TMV VIRIONS
TMV virions are rod shaped, 300nm long and about 18nm in diameter. The virions have helical symmetry and a hollow, cylindrical core.
Component
Number of molecules in virion 1 about 2140
Molecular weight
RNA [6395 residues]* Capsid protein
2.3 x 106 17,500
* The virion RNA has a 5' cap structure at the 5' nucleotide residue
Hull, p. 132
The regular icosahedron: Symmetry
12 vertices, 20 identical triangular faces 5 fold rotational symmetry center of face - 3 fold symmetry axis midpoint of each edge - 2 fold symmetry axis
Variations on a Theme Objective: Make particles larger and more spherical Strategy: Divide the original 20 faces in smaller faces, which each again can be filled with subunits Triangulation: T=Px(f)2
Many viruses: P=3, f=1, T=3; (pentamers and hexamers of subunits)
Page 136
Icosahedral Symmetry
In higher order icosahedra, the symmetry of the particle is defined by the triangulation number of the icosahedron. The triangulation number, number T, is defined by:T = f 2 x P where f is the number of subdivisions of each side of the triangular face, f 2 is the number of subtriangles on each face & P = h2 + hk + k2, where h & k are any distinct, nonnegative integers.
Fooling around with P
Fig. 5.17, page 137 Each original face is divided up in 6x1/2 new faces: P=3 Each new sub-triangle again can handle 3 protein units T=Pxf2: 3x1=3===>60x3=180 protein subunits
Triangulation Numbers
Molecular Virology, 3rd edition, Academic Press
icosahedral Cowpea mosaic (como) virus virion
Three copies of L coat protein
Five copies of S coat protein
(12 vertices) x (5 S protein per vertex) = 60 copies of S protein
(20 faces) x (3 L protein per face) = 60 copies of L protein
Capsid is composed of equal molar amounts of two coat proteins, L and S
Virus Structure
(Reconstruction based on X-Ray crystal structures)
Tomato Bushy Stunt Virus
Cowpea Chlorotic Mottle Virus
D. M. Rochon, Canada
Genome organisation
The Mimivirus has a size larger than the smallest bacteria and, with about 900 genes, a genetic complexity greater than that of the most reduced bacteria
[Link]